Functional imaging of living Paramecium by means of confocal and two-photon excitation fluorescence microscopy

Alberto Diaspro; Paola Fronte; Marco Raimondo; Marco Fato; Gianluca DeLeo; Francesco Beltrame; Fabio Cannone; Giberto Chirico; Paola Ramoino

doi:10.1117/12.468356

28 May 2002 Functional imaging of living Paramecium by means of confocal and two-photon excitation fluorescence microscopy

Alberto Diaspro, Paola Fronte, Marco Raimondo, Marco Fato, Gianluca DeLeo, Francesco Beltrame, Fabio Cannone, Giberto Chirico, Paola Ramoino

Author Affiliations +

Proceedings Volume 4622, Optical Diagnostics of Living Cells V; (2002) https://doi.org/10.1117/12.468356
Event: International Symposium on Biomedical Optics, 2002, San Jose, CA, United States

Abstract

Confocal and Two-photon excitation laser scanning microscopy allow gathering three-dimensional and temporal information from biological systems exploiting fluorescence labeling and autofluorescence properties. In this work we study biological events linked to functionality in Paramecium primaurelia. The internalization of material in ciliated one-celled organisms (protozoa) occurs via different mechanisms, even if most of nutrients, particulate or not, is taken up by food vacuoles formed at the bottom of the oral cavity. The endocytosis of small-sized molecules occurs at the parasomal sacs, located next the ciliar basal bodies. Vital fluorescent dyes (BSA-FITC, WGA-FITC, dextran-Texas Red, cholesteryl-Bodipy) and autofluorescence were used to study formation, movement, and fusion of vesicles during endocytosis and phagocytosis of Paramecium primaurelia. By immobilizing living cells pulsed with food vacuole and endosome markers at successive times after chasing in unlabeled medium, the intracellular movement and fusion of food vacuoles and of endosomes were visualized. A temporal analysis of fluorescence images and the false-color technique were used. Starting from time series or 3D data sets composite images were generated by associating with each originally acquired image a different color corresponding to each sampling point in time and along the z-axis. Second Harmonic Generation Imaging attempts are also outlined.

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Alberto Diaspro, Paola Fronte, Marco Raimondo, Marco Fato, Gianluca DeLeo, Francesco Beltrame, Fabio Cannone, Giberto Chirico, and Paola Ramoino "Functional imaging of living Paramecium by means of confocal and two-photon excitation fluorescence microscopy", Proc. SPIE 4622, Optical Diagnostics of Living Cells V, (28 May 2002); https://doi.org/10.1117/12.468356

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